Hi,
I am having trouble staining for slow vs fast muscle fibers on 4% PFA fixed mouse calf cross sections. Currently I am trying the Vector M.O.M kit with either:
Abcam Anti-Slow Skeletal Myosin Heavy chain antibody [NOQ7.5.4D]
Abcam Anti-Fast Myosin Skeletal Heavy chain antibody [MY-32]
And detecting with DAB staining. The result is not ideal and seems to vary between tissue.
What is the most optimal procedure for staining general fast vs slow fibers in perfused mouse tissue? Are non-IHC protocols preferable for PFA fixed tissue? The tissue has already been collected for all animals and fixed in PFA, therefore fresh tissue is currently not an option.
I was having issues with muscle fiber typing and posted a similar question a few years back. Research gate user Stefano Ciciliot, was incredibly helpful in that endeavor. I'd suggest checking him out and his responses to other questions. He was a lifesaver. Good luck!
I am having the same issue now... did you ever figure anything out to use?
In my experience so far, the slow antibody mentioned above does seem to work in perfused muscle tissue (not as well as fresh tissue) with the help of antigen retrieval by boiling sections in sodium citrate buffer, however the fast does not seem to work no matter the treatment. Seems like its better not to perfuse the tissue in the first place and leave it as fresh frozen.
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